Changes in the intracellular Ca²⁺ content in human red blood cells in the presence of glycerol

Changes of the intracellular Ca²⁺ content in human red blood cells (RBCs) in glycerol-containing solutions and after freeze-thawing the cells with glycerol and subsequent deglycerolization were investigated with the Ca²⁺-sensitive fluorescent dye fluo-4 using fluorescence microscopy. In the glycerol-containing solutions the Ca²⁺ content increased when compared with a physiological medium (Hepes buffered saline solution (HBSS)). This effect was most likely a result of an inhibition of the Ca²⁺ pump. After inhibiting the Ca²⁺ pump using o-vanadate, the Ca²⁺ uptake was not significantly different in the cells in glycerol-containing and physiological medium. Freeze-thawing and deglycerolization of RBCs resulted in a more pronounced increase in the Ca²⁺ content. Also in this case, the Ca²⁺ pump seemed to play a major role.